Mannosylated PLP139–151 induces peptide-specific tolerance to experimental autoimmune encephalomyelitis
Introduction
C-type lectins play an important role in the recognition of carbohydrate structures on pathogens and endogenous ligands. Members of this family play an important role in immune regulation, immune evasion, and tolerance (Figdor et al., 2002, van Kooyk and Geijtenbeek, 2003, Steinman et al., 2003). One of these members is the mannose receptor that selectively binds carbohydrate groups such as mannose and fucose on glycoproteins of yeast, bacteria, parasites, and certain viruses and is expressed on subsets of antigen-presenting cells (Stahl and Ezekowitz, 1998, Sallusto et al., 1995). Binding of mannosylated ligands leads to internalization and delivery to the late endosomal compartments where low pH releases the ligand from the receptor, after which the latter is recycled to the cell surface (Koning, 1997, Tan et al., 1997, Tan et al., 1998). Mannose receptor-mediated endocytosis is a very efficient process allowing multiple rounds of antigen binding and endosomal delivery and can result in up to a 10,000-fold increased efficiency of presentation of mannosylated antigen compared to non-mannosylated antigen (Engering et al., 1997, van Bergen et al., 1999). Another member of the family of C-type lectins is DEC-205, a multilectin receptor homologous to the macrophage mannose receptor that is selectively expressed by immature DC (Mahnke et al., 2000). Targeting of ovalbumin to this receptor was shown to induce T cell unresponsiveness unless a second activation signal was provided simultaneously (Hawiger et al., 2001, Bonifaz et al., 2002). Interestingly, binding of the mycobacterial cell wall component ManLAM to the DC-specific C-type lectin DC-SIGN inhibits DC maturation and induces IL-10 production (Geijtenbeek et al., 2003). Consequently, a role of C-type lectins in the maintenance of tolerance for self-antigens has been suggested (van Kooyk and Geijtenbeek, 2003, Steinman et al., 2003). To address this possibility we studied a mannosylated self-peptide with regard to its ability to induce disease and show for the first time that a mannosylated form of PLP139–151 (M-PLP139–151) induces a state of tolerance to EAE.
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Synthesis of peptides
Mannosylation of PLP139–151 was performed as described elsewhere (Tan et al., 1997, van Bergen et al., 1999). Briefly, PLP139–151 was synthesized using solid phase synthesis. For M-PLP139–151 the peptide was N-terminally elongated with a lysine building block containing two tetra-acetyl protected mannose groups. The PLP139–151 peptide was elongated with bis-acetyl lysine. Cleavage of the peptides and removal of the protecting groups was performed with TFA. Removal of the acetyl protecting
Mannosylated PLP139–151 induces tolerance to EAE
To evaluate if targeting of C-type lectins would induce tolerance to EAE we first compared encephalitogenic PLP139–151 and its mannosylated form (M-PLP139–151) with respect to their efficacy to induce EAE when emulsified in complete adjuvant containing M. tuberculosis H37RA.
To this end we immunized groups of mice subcutaneously with 0.5, 5, or 50 μg of either peptide emulsified in complete adjuvant. As shown in Fig. 1, the severity of EAE during the first 4 weeks after induction was dependent
Discussion
C-type lectins have gained a lot of interest in view of their role as receptors that facilitate the interaction between DC and microorganisms and the subsequent induction of a protective immune response. In contrast, it has also been demonstrated that such interactions may result in the induction of tolerance. Targeting of ovalbumin to DEC205, a mannose-receptor expressed on immature DC, induces unresponsiveness in vivo (Hawiger et al., 2001, Bonifaz et al., 2002). However, as yet, no data are
Acknowledgements
We are grateful to Joost Drexhage, Inge Haspels, Bep Blauw, and Willemien Benckhuijsen for excellent technical assistance. These studies were supported by grants 98-366 and 00-432 from The Dutch Foundation for MS Research.
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